Preparation of 7-halo-6-deoxy-tetracyclines



United States Patent This invention relates to the preparation of7-halo-6- deoxytetracyclines which may be represented by the followinggeneral formula 112. H R2 R3 (C s)2 -CONH2 (at E ii wherein R is Br, C1or I and R is H or CH when R, is

H, and R is CH when R is OH.

The halodeoxytetracyclines of this invention may be prepared bydissolving asuitable 6-deoxytetracycline, i.e. 6-deoxytetracyclineitself, 6-demethyl-fi-deoxytetracycline or5-hydroxy-o-deoxytetracycline, in a suitable solvent,

such as concentrated mineral acid, i.e., hydrochloric acid, sulfuricacid, etc. and adding a halogenating agent such as an N-haloamide, i.e.,N-bromosuccinimide, N-bromoacetamide, N-bromophthalimide,N-chlorosuccinimide, N- iodosuccinimide, etc. The reaction with thehaloamide is preferably carried out at temperatures of from about 0 C.to about 20 C. until the reaction is complete. Alternatively, thebromination may be carried out with bromine in a strong mineral acid,i.e., HBr and in a suitable organic solvent such as a lower alkanoicacid, i.e. acetic acid, propionic acid, etc, at temperatures of fromabout 10 C. to about 50 C. The halodeoxytetracycline so-formed isisolated from the reaction mixture by any convenient method as, forexample, by precipitation with ethyl ether or the like, and the productmay be purified by recrystallization from an alcohol-acetone-ethersolution in a standard manner.

The halodeoxytetracyclines are biologically active and have thebroad-spectrum antibacterial activity of the previously knowntetracyclines. The antibacterial spectrum of certain of these compounds,representing the amount required to inhibit the growth of varioustypical bacteria, was determined in a standard manner by the agardilution streak technique which is commonly used in testing newantibiotics. The minimal inhibitory concentrations, expressed in gamrnasper milliliter, of 7- bromo-o-demethyl-6-deoxytetracycline, 7bromo-6-deoxytetracycline and 7-iodo 6 deoxy-tetracycline againstvarious test organisms are reported in the table below. For comparisonpurposes the antibacterial activity of tetracycline against the sameorganisms is also included.

TABLE 7-Bromo-6- 7-Bromo-6- 7-I0d0- Organism Tetra-Deoxytetradernethyl-6- 6-dcoxycycline cycline deoxytctratetracyclinecycline Mycobacterium rauae 1 0. 5 0.25 4 lllycobacteriurn smeumatisATOC 607 1 l 0. 25 8 Staphylococcus aureus 209P. 2 1 1 2 Sarciua luteo1001 2 1 0.5 2 Bacillus subtilis AICO 6633- 0. 5 0. 25 0. 25 0. 5Streptococcus pyogeues C203. 0. 5 0. 25 0. 5 2 Streptococcus 'y N o.11..- 250 4 4 4 Staphylococcus albus N o. 69. 250 8 8 15 Streptococcus BN o. 250 4 2 4 Staphylococcus aureus NY 104 2 1 1 2 Bacillus cereus N o.0. 25 0.25 0. 5 2 Pseudomouas acruginosa 15 250 31 250 Proteus vulaaris8427 15 8 2 31 Escherichia coli ATCC 9637- 15 250 31 250 Salmonellagallinarum 8 250 31 250 Escherichia coli No. 22 4 15 8 62 The7-iododeoxytetracycline and 7-bromodeoxytetracycline are particularlyuseful as diagnostic agents in the detection of cancer. The7-iododeoxytetracycline and 7- bromodeoxytetracycline appear toconcentrate in rapidly proliferating tissue such as found in tumorgrowths. These compounds may be rendered radioactive by incorporation ofiodine 131 or bromine 82 in the 7-position of the ring nucleus. Theseradio labeled compounds by virtue of their gamma rays and beta particleemissions allow the detection, localization and diagnosis of neoplastictissue. The chemical methods for preparing these radio-labeled compoundsare similar to those preparative methods already described for thepreparation of non-radioactive compounds with the obvious exception thatthe radioisotope of the appropriate halogens are used.

The invention will be described in greater detail in conjunction withthe following specific examples.

Example 1 PREPARATION OF 7-BROMO-G-DEOXYTETRACYCLINE To a solution of0.2 gram of 6-deoxytetracycline [J.A.C.S. 80, 5324 (1958)] in 10milliliters of concentrated sulfuric acid at 0 C. is added 76 milligramsof N-bromosuccinimide. The reaction mixture is kept at 0 C. for 30minutes, and then added slowly drop-wise to 200 milliliters of coldether. A solid precipitates which is filtered and dried. The product.weighs 160 milligrams. This product is crystallized fromethanol-acetone-ether solution to give milligrams of purified product.This product is 1.21.5 times as active biologically as tetracycline.

Example 2 PREPARATION OF 7-BROMO-6-DEMETHYL-6-DEOXY- TETRACYCLINE Asolution of 200 milligrams (0.045 m mole) of 6-demethyl-6-deoxytetracycline hydrochloride [J.A.C.S. 80, 5 324 (1958)]and 80 milligrams (0.45 m mole) of N- bromosuccinimide in 5.0milliliters of concentrated sulfuric acid is stored at ice-bathtemperature for 30 minutes. The reaction mixture is slowly added to 250milliliters of cold ether and the solid that separates weighs 0.2 gram.A portion (25 milligrams) of this material is recrystallized fromethanol/ether yielding 19 milligrams of pure7-brorno-6-demethyl-6-deoxytetracycline which is 2.2 to 2.5 times asactive as tetracycline.

Example 3 PREPARATION OF 7-CHLORO-6-DEOXYTETRACYCLINE The procedure ofExample 1 is followed except that N-chlorosuccinirnide is used as thehalogenating agent. The product is isolated as in Example 1 and7-chloro-6- deoxyltetracycline is obtained.

Example 4 PREPARATION OF 7-CHLORO-6-DEMETHYL-6-DEOXY- TETRACYCLINE Theprocedure of Example 2 is followed except that N-chlorosuccinimide isused as the halogenating agent. The product is isolated as in Example 2and 7-chloro-6- demethyl-6-deoxytetracycline is obtained.

Example 5 PREPARATION OF 7-BROM0-6-DEOXYTETRACYCLINE 85.6 milligrams of6-deoxytetracycline is dissolved in 3.9 milliliters of acetic acid and 1milliliter of 30% HBracetic acid is added. With stirring, 0.22milliliter of 1 molar bromine in acetic acid is added excess of 1equivalent). After standing about 65 hours at room temperature, 10milliliters of ether are added and a yellow crystalline solid slowlydeposits. After 6 hours the solid is filtered off and dried; weight,68.4 milligrams.

Example 6 PREPARATION OF 7-BROMO-5-HYDROXY-6-DEOXY TETRACYCLINE Asolution of 200 milligrams (0.42 m mole) ofS-hydroxy-6-deoxytetracycline hydrochloride [J.A.C.S. 80, 5324 (1958)]and 75 milligrams (0.42 m mole) of N- bromosuccinimide in 5.0milliliters of concentrated sulfuric acid is stirred at ice-bathtemperature for 10 minutes. The reaction solution is slowly poured into250 milliliters of cold ether. The solid is filtered and dried; yield1.8 grams. This material is converted to the free base by dissolving inwater, adjusting the pH of the solution to 6.5 with 1 N sodium carbonateand extraction with n-butanol; yield 75 milligrams.

Microbiological activity equals 45% of tetracycline (activity ofstarting S-hydroxy-6-deoxytetracycline equals 30% of tetracycline).

Example 7 PREPARATION OF 7-IODO-6-DEOXYTETRACYCLINE SULFATE A solutionof 200 milligrams (0.38 m mole) of 6-deoxytetracycline sulfate and 85.5milligrams (0.35 m mole) of N-iodosuccinimide in 5.0 milliliters ofconcentrated sulfuric acid is stirred at 0 C. for forty minutes. Themixture is added dropwise to 250 milliliters of cold ether. The sOlidthat separates Weighs 0.16 grams. A portion (50 milligrams) of thismaterial is recrystallized from methyl cellosolve/chloroform; yield 22milligrams.

Example 8 PREPARATION OF 7-IODO-5-HYDROXY-6-DEOXYTETRA- CYCLINE SULFATEThe procedure of the preceding example is following except that5-hydroxy-6-deoxytetracycline is used. 7-iodo-5-hydroxy-6-deoxytetracycline is obtained.

Example 9 7-10DO-6-DEMETHYL-6-DEOXY'IETRACYCLINE SULFATE To a solutionof 0.2 gram (0.39 m mole) of 6-demethyl-6-deoxytetracycline sulfate in5.0 milliliters of cold (0 C.) concentrated sulfuric acid is added 88milligrams (0.39 m mole) of N-iodosuccinimide. The mixture is stored at0 C. for forty minutes and slowly poured 7 -halo-6-deoxytetra- 'M H a as)2 CONH2 l H OH O OH O wherein R is a member of the group consisting ofbromine, chlorine and iodine and R is a member of the group consistingof hydrogen and methyl when R is hydrogen and R is methyl when R ishydroxy which comprises treating a compound of the group consisting of6-deoxytetracycline, 6-demethyl-6-deoxytetracycline and5-hydroxy-6-deoxytetracycline with an N-haloamide of the groupconsisting of N-bromosuccinimide, N-bromoacetamide, N-bromophthalirnide,N-chlorosuccinimide, and N-iodosuccinimide in the presence of aconcentrated mineral acid at a temperature of from about 0 C. to about20 C.

2. The method of preparing 7-bromo-6-deoxytetra cycline which comprisestreating 6-deoxytetracyc1ine with N-bromosuccinimide in the presence ofa concentrated mineral acid at a temperature of from about 0 C. to about20 C.

3. The method of preparing 7-bromo-6-demethyl-6-deoxytetracycline whichcomprises treating 6-demethyl-6- deoxytetracycline withN-bromosuccinimide in the presence of a concentrated mineral acid at atemperature of from about 0 C. to about 20 C.

4, The method of preparing 7-chloro-6-deoxytetracycline which comprisestreating 6-deoxytetracycline with N-chlorosuccinimide in the presence ofa concentrated mineral acid at a temperature of from about 0 C. to about20 C.

5. The method of preparing 7-chloro-6-demethyl-6-deoxytetracycline whichcomprises treating 6-demethyl-6- deoxytetracycline withN-chlorosuccinimide in the presence of a concentrated mineral acid at atemperature of from about 0 C. to about 20 C.

6. The method of preparing 7-bromo-5-hydr0xy-6-deoxytetracycline whichcomprises treating 5-hydroxy-6-deoxytetracycline withN-bromosuccinirnide in the presence of a concentrated mineral acid at atemperature of from about 0 C. to about 20 C.

7. The method of preparing 7-iodo-6-deoxytetracycline which comprisestreating 6- deoxytetracycline with N-iodosuccinimide in the presence ofa concentrated mineral acid at a temperature of from about 0 C. to about20 C.

8. The method of preparing 7-iodo-6-demethyl-6-deoxytetracycline whichcomprises treating 6-demethy1-6- deoxytetracycline withN-iodosuccinimide in the presence of a concentrated mineral acid at atemperature of from about 0 C. to about 20 C.

9. The method of preparing 7-iodo-5-hydroxy-6-deoxytetracycline whichcomprises treating 5-hydroxy-6-deoxytetracycline with N-iodosuccinimidein the presence of a concentrated mineral acid at a temperature of fromabout 0 C. to about 20 C.

wherein R is a member of the group consisting of hydrogen and methylwhen R is hydrogen and R is methyl when R is hydroxy which comprisesreacting a compound of the group consisting of 6-deoxytetracyc1ine, 6-demethyl-6-deoxytetracycline and S-hydroxy 6 deoxytetracycline withbromine in a strong mineral acid, and in the presence of an organicsolvent at a temperature of from about 10 C. to about 50 C.

11. The method of preparing 7-bromo-6-doxytetracycline which comprisesreacting 6-deoxytetracycline with bromine in a strong mineral acid andin the presence of an organic solvent at a temperature of from about 10C. to about 50 C.

References Cited in the file of this patent UNITED STATES PATENTS RitterFeb. 28, 1956 OTHER REFERENCES Geschickter: J. Am. Med. Assoc., Feb. '1,1930, pages 326328.

Fieser et 211.: Natural Products Related to Phenanthrene, pages 388-389,458; 533-536. Reinhold Pub. Co., New York (1949).

Abbott Laboratories, Diagnostic Procedures With Radioisotopes, pages 23,28 (January 1955).

Migrdichian: Organic Synthesis, vol. II, page 924, Reinhold Pub. Co.,New York (1957).

Rall et al.: Journ. US. National Institute of Cancer,

20 pages 79-83 (July 1957).

Stephens et al.: I. Am. Chem. Soc., vol. 80, pages 5324-5 (October1958).

1. THE METHOD OF PREPARING 7-HALO-6-DEOXYTETRACYCLINES OF THE FORMULA:WHEREIN R1 IS A MEMBER OF THE GROUP CONSISTING OF BROMINE, CHLORINE ANDIODINE AND R2 IS A MEMBER OF THE GROUP CONSISTING OF HYDROGEN AND METHYLWHEN R3 IS HYDROGEN AND R2 IS METHYL WHEN R3 IS HYDROXY WHICH COMPRISESTREATING A COMPOUND OF THE GROUP CONSISTING OF 6-DEOXYTETRACYCLINE,6-DEMETHYL-6-DEOXYTETRACYCLINE AND 5-HYDROXY-6-DEOXYTETRACYCLINE WITH ANN-HALOAMIDE OF THE GROUP CONSISTING OF N-BROMOSUCCINIMIDE,N-BROMOACETAMIDE, N-BROMOPHTHALIMIDE, N-CHLOROSUCCINIMIDE, ANDN-IODOSUCCINIMIDE IN THE PRESENCE OF A CONCENTRATED MINERAL ACID AT ATEMPERATURE OF FROM ABOUT 0*C. TO ABOUT 20*C.